This project advances a new therapeutic paradigm of selective, pharmacological control of microbiome function to improve human health. The use of non-steroidal anti-inflammatory drugs (NSAIDs) is associated with intestinal ulceration, inflammation and bleeding that result in reduced quality of life, hospitalization, and even death. These NSAID-induced side effects are triggered by an enzyme in the intestinal microbiome-the bacterial ?-glucuronidase (bGUS) enzyme, whose activity results in the production of toxic NSAID by-products that damage the intestinal lumen. This proposal aims to synthesize and characterize new drug candidates that selectively inhibit bacterial bGUS to be developed as therapeutic adjuncts to improve the benefit-to-risk outcomes associated with NSAID therapy. We previously reported the discovery of the first class of compounds that selectively blocks bacterial bGUS without exhibiting antibiotic activity. As a proof-of-concept, a prototype bGUS inhibitor (Inh-1) was shown to alleviate all measures of enteropathy in mice induced by three NSAIDs: diclofenac, indomethacin, and ketoprofen. We have since identified another non-proprietary, selective, and non-toxic bacterial bGUS inhibitor (Inh-20) that is >100-fold more potent than Inh-1. Inh-20 is a promising scaffold for hit-to-lead chemistry and lead optimization. However, Inh-20 undergoes extensive first- pass metabolism and intestinal absorption following oral dosing. We propose to rationally design and synthesize novel analogues that retain the potency, selectivity and non-antibiotic activity of Inh-20, while exhibiting low intrinsic metabolim and oral bioavailability. The proposed analogues are expected to be more efficacious in the GI tract and less likely to produce systemic side effects. This project will employ proven drug discovery approaches to enable the identification of a promising, novel lead series that can be further optimize into a clinical candidate. Our research strategy lays the groundwork for the discovery of the first microbiome- targeted, human therapeutic that is: (1) Effective in reducing the risks of lower GI damage from NSAID use; (2) not harmful to beneficial intestinal microbes; and (3) safe for the human host.